Laser Optoacoustic Ultrasonic Imaging System (LOUIS) and Methods of Use

ABSTRACT

Provided herein are the systems, methods, components for a three-dimensional tomography system. The system is a dual-modality imaging system that incorporates a laser ultrasonic system and a laser optoacoustic system. The dual-modality imaging system generates tomographic images of a volume of interest in a subject body based on speed of sound, ultrasound attenuation and/or ultrasound backscattering and for generating optoacoustic tomographic images of distribution of the optical absorption coefficient in the subject body based on absorbed optical energy density or various quantitative parameters derivable therefrom. Also provided is a method for increasing contrast, resolution and accuracy of quantitative information obtained within a subject utilizing the dual-modality imaging system. The method comprises producing an image of an outline boundary of a volume of interest and generating spatially or temporally coregistered images based on speed of sound and/or ultrasonic attenuation and on absorbed optical energy within the outlined volume.

CROSS-REFERENCE TO RELATED APPLICATIONS

This is a continuation under 35 U.S.C. § 120 of pending non-provisionalapplication U.S. Ser. No. 13/748,498, filed Jan. 23, 2013, which claimsbenefit of priority under 35 U.S.C. § 119(e) of provisional applicationU.S. Ser. No. 61/605,276, filed Mar. 1, 2012, now abandoned and ofprovisional application U.S. Ser. No. 61/632,387, filed Jan. 23, 2012,now abandoned, all of which are hereby incorporated by reference.

BACKGROUND OF THE INVENTION Field of the Invention

The present invention relates to the field of biomedical imaging anddiscloses the designs and methods used for a tomographic system that canprovide comprehensive medical information about a portion of the bodyunder examination. More specifically, the present invention provides aLaser Optoacoustic Ultrasonic Imaging System (LOUIS) forthree-dimensional tomography of a subject or portion or body partthereof.

Description of the Related Art

Imaging internal structures of a human or animal subject body has been asubject of many inventions. There are systems that use ultrasoundpressure waves, photon waves and acoustic waves induced by absorption ofphotons in tissues of the subject body. However, the prior art lacks asystem that can provide comprehensive information about tissues,including anatomical structure (morphology) and molecular compositionsimultaneously with information about tissue normal or abnormalfunction. The most detailed and comprehensive information can beprovided by high resolution three dimensional maps, especially valuableif such maps are provided in real time, i.e. faster than the timerequired for certain changes to occur in the subject body. Medicallyimportant changes may occur in the subject body on the time scale aslong several minutes and as short as fraction of a second. Therefore,the most ideal system can provide detailed (high resolution)three-dimensional functional and anatomical maps (images) of the subjectbody or least certain organs of the subject body.

Laser ultrasound method and systems designed for nondestructiveevaluation of materials such as metals, ceramics and fiber-epoxycomposites have been discussed in the literature. However, these systemsare not three-dimensional tomography systems and their design cannot beused for biomedical imaging. Methods and materials for laser generationof ultrasonic pulses have been discussed in the prior art (7) andproposals have been made by O'Donnell group for application of suchpulses in 3D and 2D ultrasonic imaging in medicine. However, the priorart lacks description of a design for 3D laser ultrasound tomographysystem capable of volumetric visualization of biomedical objects throughalgorithms of reconstruction tomography, such as filteredback-projection tomography and of the full set of properties of thelayers of the materials for the most effective generation ofultrawide-band ultrasound with laser pulses. Three-dimensionalultrasound tomography has been proposed for biomedical imaging,specifically for the volumetric imaging of breast cancer. However, theultrasound pulses in these systems are generated through application ofelectrical voltage pulses to piezoelectric elements.

Optoacoustic tomography is used in biomedical applications for in vivoand in vitro imaging of animal and human tissues and organs based ondifferences in tissue optical properties. Optoacoustic tomography hasthe potential to become valuable modality of functional molecularimaging. The essence of functional molecular imaging is to providequantitative information (maps) of distributions and concentrations ofvarious molecules of interest for medicine. For example, distribution ofhemoglobin and oxi-hemoglobin concentration in tissue shows whether thetissue normally functions or it is damaged or malignant. Distribution ofspecific protein receptors in cell membranes give insight into molecularbiology or cells helping in designing drugs and therapeutic methods totreating human diseases.

Laser optoacoustic imaging systems and methods have been disclosed byOraevsky et al (8,9), Kruger et al. (10-11) and others (12-18). However,the prior art lacks description of a 3D tomography system that combineslaser ultrasonic and laser optoacoustic tomography in one imagingmodule, allows natural coregistration of volumetric images acquired andreconstructed using the two modalities and thereby provides the mostcomprehensive anatomical, functional and molecular information for thephysician or biomedical researcher.

The prior art contains some limited information about the idea ofcombining the laser optoacoustic imaging and the laser ultrasonicimaging. Specifically, the group of Karabutov from Moscow StateUniversity proposed a combined array that can be used in both imagingmodalities. However, the proposed design was limited to a scanningsystem based on a single transducer that is focused into a point at somespecific depth (19). This design could only be used for one-dimensionaldepth profiling, potentially for two-dimensional imaging, even thoughthe design is shown only for a single transducer, but not forthree-dimensional tomography. This design remains just an idea severalyears after the original publication likely because authors themselvesrealized a number of technical deficiencies limiting usefulness of thissystem in biomedical applications.

A major drawback of this design is that the array is focused into a lineand it will take a long time to acquire complete 2D image of a slice,which is not practical. Moreover, the main problem in the design is thatit cannot be used for optoacoustic imaging as described because thelaser pulse strikes a strongly absorbing polymer layer and there is nolaser pulse delivery directly to the tissue surface. Therefore, eventhough the paper implies a combined laser ultrasound and optoacousticsystem, the proposed array can be used only for laser ultrasound imagingwhich is similar to the designs developed for laser ultrasoundnondestructive evaluation of industrial materials.

Despite years of research effort, there remains an urgent need for thedevelopment of imaging technology that can improve the sensitivity ofdetection, specificity of biomedical diagnostics and characterization ofchanges that occur during and after therapeutic interventions byproviding comprehensive detailed unobstructed high resolution volumetricpictures of biological tissues, organs and bodies. Detection andtreatment of breast cancer especially is lacking the neededtechnologies. The current problems of breast cancer care are numerous(1-5), i.e., a large number (˜20%) of breast tumors are missed by x-raymammography, especially in the dense breast of younger women, (2) about75% of biopsies are unnecessary, cancers are missed due to insufficientcontrast of ultrasound guided biopsy, and a lack of fast and safefunctional imaging techniques to assess the effectiveness of anticancerchemotherapy and other therapies. Diagnostic and treatment of many otherdiseases (atherosclerosis and peripheral vascular diseases, heartdisease and stroke, diabetes and burns) and biomedical research (incancer biology, hematology, neurology and drug discovery and testing)can benefit from the comprehensive 3D tomography system.

While prior art systems may provide a base for the design anddevelopment of a clinically viable laser optoacoustic ultrasonic imagingsystem (LOUIS) (19,20), Previously developed optoacoustic imagingsystems and laser ultrasound monitoring systems have limited resolutionand sensitivity, have limited field of view, have reduced accuracy ofquantitative information, have artifacts associated with projection ontoimage plane of objects located out of the image plane, and have nocapability to provide detailed information on distribution of speed ofsound.

Thus, there is a recognized need in the art for an improvedthree-dimensional tomographic system that overcome these limitations.Particularly, the prior art is deficient in a tomographic system thatcombines laser ultrasound and laser optoacoustic tomography useful formany biomedical applications such as, but not limited to, cancerdetection or screening, monitoring of anticancer therapies, detectionand characterization of vascular diseases, monitoring drug distribution,distribution of nanoparticles or contrast agents and physiological andpathological processes. The present invention fulfills this longstandingneed and desire in the art.

SUMMARY OF THE INVENTION

The present invention is directed to a laser ultrasonic imaging system.The imaging system comprises means for delivering short pulses ofoptical energy to an array of ultrasonic emitters comprising opticallyabsorbing elements placed in specific locations configured for efficientconversion of the absorbed optical energy into a short pulses ofacoustic energy within a wide band of ultrasonic frequencies. Theimaging system comprises means for delivering the short ultrasonicpulses with known amplitude and ultrasonic frequency spectrum through acoupling medium to a volume of interest in a subject at a given time ortime zero. The imaging system comprises means for detecting theultrasonic pulses in multiple positions at or around said volume ofinterest and measuring one or more parameters of time of propagation,amplitude and ultrasonic frequency spectrum, after the ultrasonic pulsesare transmitted through or reflected from the volume of interest usingan array of wide-band ultrasonic transducers that convert ultrasonicpulses into electronic signals. The imaging system comprises means foranalog amplification and digital recording of the electronic signals andfor performing signal processing to remove distortions of electronicsignals. The imaging system comprises means for image reconstructionusing mathematical tomography algorithms, means for image processing anddisplay and for data transmission and system control.

The present invention also is directed to a dual-modality imagingsystem. The dual-modality imaging system comprises a first meanscomprising the laser ultrasonic system described herein configured togenerate tomographic images of a volume of interest in a subject bodyutilizing parameters comprising one or more of the speed of sound,ultrasound attenuation or ultrasound backscattering. The dual-modalityimaging system comprises a second means for generating optoacoustictomographic images of distribution of the optical absorption coefficientin the subject body utilizing parameters of the absorbed optical energydensity or various quantitative parameters that can be derived from theoptical absorption.

The present invention is directed further to a imaging method forincreasing contrast, resolution and accuracy of quantitative informationobtained within a subject. The method comprises the steps of producing alaser ultrasound or laser optoacoustic image of an outline boundary of avolume of interest within the subject using the dual-modality imagingsystem described herein. A spatially or temporally coregistered image ofspeed of sound and/or an image of ultrasonic attenuation within theoutlined volume boundary is generated from information contained in thelaser ultrasound or laser optoacoustic image and a spatially ortemporally coregistered optoacoustic image is generated based onabsorbed optical energy using an algorithm of the image reconstructionthat employs distribution of the speed of sound and/or ultrasoundattenuation within the outlined volume boundary.

The present invention is directed further still to a laser optoacousticultrasound imaging system (LOUIS). The LOUIS imaging system comprises adual laser source switchable between a laser ultrasonic mode and a laseroptoacoustic mode, where the laser source is configured to emit eithershort optical pulses with high repetition rate for the illumination ofthe ultrasonic emitters in the ultrasonic mode or short optical pulseswith lower repetition rate but higher pulse energy for the illuminationof the volume of interest in the optoacoustic mode. The LOUIS imagingsystem comprises an imaging module comprising one or more ultrawide-bandultrasonic transducers configured to detect, through a coupling medium,optoacoustic and ultrasonic signals propagated as transient pressurewaves from the volume of interest within a subject body. The LOUISimaging system comprises means to rotate and/or translate the imagingmodule relative to the volume of interest in the subject body to createmultiple pressure waves, said means computer controllable or manuallycontrollable. The LOUIS imaging system comprises means for processingthe detected laser optoacoustic and laser ultrasonic signals and forreconstructing processed signals into one or more of anatomical andfunctional/molecular images of the volume of interest in the subjectbody. The present invention is directed to a related LOUIS imagingsystem further comprising means for displaying the one or more images orsuperimposed coregistered images of the subject body or the volume ofinterest therein.

The present invention is directed further still to a method for imaginga subject's body or a volume of interest therewithin. The methodcomprises positioning the subject body within or proximate to theimaging module of the laser optoacoustic ultrasound imaging systemdescribed herein, delivering a laser-generated pulses of ultrasonicenergy to a volume of interest in the subject body and detecting thetransmitted or reflected ultrasonic pressure waves while measuring oneor more parameters comprising a difference between the time of emissionand a time of arrival, a difference between emitted amplitude anddetected amplitude, and a difference between ultrasonic frequencyspectrum of emitted and detected ultrasonic pulses. Then delivering alaser-generated pulse of optical energy is delivered to a volume ofinterest in the subject body and the ultrasonic pressure waves generatedthrough optical absorption inside the subject body are detected whilemeasuring one or more parameters comprising a time of arrival relativeto a time of generation, an amplitude of detected optoacoustic signals,and an ultrasonic frequency spectrum of detected optoacoustic signals.The subject body or volume of interest therein is scanned with adetecting array of ultrawide-band ultrasonic transducers by repeatingsteps the previous steps at multiple positions around the subject bodyor volume of interest while simultaneously scanning the sources ofoptical energy and sources of ultrasonic energy such that relativeposition of the detecting array of ultrasonic transducers and thesources of optical or ultrasonic energy can change or remain constantduring the scans. processing the detected ultrasonic signals areprocessed to remove distortions of detected signals and one or morevolumetric images are reconstructed via mathematical tomographyalgorithms using data of the processed signals.

Other and further aspects, features, and advantages of the presentinvention will be apparent from the following description of thepresently preferred embodiments of the invention given for the purposeof disclosure.

BRIEF DESCRIPTIONS OF THE DRAWINGS

So that the matter in which the above-recited features, advantages andobjects of the invention, as well as others that will become clear, areattained and can be understood in detail, more particular descriptionsof the invention briefly summarized above may be had by reference tocertain embodiments thereof that are illustrated in the appendeddrawings. These drawings form a part of the specification. It is to benoted, however, that the appended drawings illustrate preferredembodiments of the invention and therefore are not to be consideredlimiting in their scope.

FIGS. 1A-1C depict two-dimensional images of a female's right cancerousbreast in an an ultrasound image (FIG. 1A), an optoacoustic image (FIG.1B) and an x-ray mammogram (FIG. 1C).

FIG. 2 shows the assembled laser optoacoustic ultrasonic system.

FIGS. 3A-3B depict the imaging module for the three-dimensional LaserOptoacoustic Ultrasound System (LOUIS-3D) with combined linear-flat plusarc shaped transducers (FIG. 3A) and with an arc-shaped transducer array(FIG. 3B).

FIGS. 4A-4C are back, front and side views, respectively, of a laserultrasonic emitter.

FIGS. 5A-5C depict the generation of Delta ultrasound pulses with highamplitude (FIG. 5A), ultrawide frequency spectrum (FIG. 5B) and widedirectivity (FIG. 5C).

FIG. 6 is a table of Gruneisen parameters for liquids and solids withhigh thermal expansion and high speed of sound.

FIG. 7 depicts a hand-held probe comprising the imaging module.

FIGS. 8A-8C are graphs of an electrically generated (FIG. 8A) and lasergenerated (FIG. 8B) ultrasound pulses and of the frequency spectrum(FIG. 8C) corresponding to FIG. 8B.

FIGS. 9A-9B depict three intersecting horse hairs (FIG. 9A) and theoptoacoustic image brightness cross-section of one hair (FIG. 9B).

FIGS. 10A-10B depict optoacoustic profiles of a PZT (FIG. 10A) and of asingle crystal PMN ceramic (FIG. 10B) ultrasonic transducers.

FIGS. 11A-11B are 2D projections of three-dimensional optoacousticimages of a mouse skin outline in vivo.

FIGS. 12A-12B illustrate the distribution of the speed of sound (FIG.12A) and ultrasonic attenuation (FIG. 12B) in a phantom simulating abreast.

FIG. 13 is a 2D projection of an optoacoustic image of mouse body.

FIG. 142D projection of a 3D LOUIS image of an animal body vasculature.

FIG. 15 is an optoacoustic image of brain vasculature in a live mouse.

FIGS. 16A-16C show 2D projections of 3D optoacoustic images usingcontrast agents of a breast tumor (FIG. 16A) before (FIG. 16B) and afterinjection of GNR-PEG-Herceptin (FIG. 16C).

FIGS. 17A-17B are 3D laser optoacoustic images of breasts acquired andreconstructed with LOUIS-3D.

FIG. 18 illustrates the optoacoustic image reconstruction algorithm.

FIGS. 19A-19B are optoacoustic images of a mouse vasculaturereconstructed with a standard filtered backprojection algorithm (FIG.19A) and with a filtered backprojection algorithm (FIG. 19B) as detailedin FIG. 18.

FIGS. 20A-20B are images reconstructed using a filtered backprojectionalgorithm and the entire set of measured signal data (FIG. 20A) andusing an iterative algorithm taking only ¼ portion of the data set (FIG.20B).

DETAILED DESCRIPTION OF THE INVENTION

As used herein in the specification, “a” or “an” may mean one or more.As used herein in the claim(s), when used in conjunction with the word“comprising”, the words “a” or “an” may mean one or more than one.

As used herein “another” or “other” may mean at least a second or moreof the same or different claim element or components thereof. Similarly,the word “or” is intended to include “and” unless the context clearlyindicates otherwise. “Comprise” means “include.”

As used herein, the term “about” refers to a numeric value, including,for example, whole numbers, fractions, and percentages, whether or notexplicitly indicated. The term “about” generally refers to a range ofnumerical values (e.g., +1-5-10% of the recited value) that one ofordinary skill in the art would consider equivalent to the recited value(e.g., having the same function or result). In some instances, the term“about” may include numerical values that are rounded to the nearestsignificant figure.

As used herein, the term “computer” or “computer system” refers to anynetworkable tabletop or handheld electronic device comprising a memory,a processor, a display and at least one wired or wireless networkconnection. As is known in the art, the processor is configured toexecute instructions comprising any software programs or applications orprocesses tangibly stored in computer memory or tangibly stored in anyknown computer-readable medium.

As used herein, the term “subject” refers to a human or other mammal oranimal or to any portion or body part thereof on which imaging, forexample, laser optoacoustic ultrasound imaging, may be performed.

In one embodiment of the present invention there is provided a laserultrasonic imaging system, comprising a) means for delivering shortpulses of optical energy to an array of ultrasonic emitters comprisingoptically absorbing elements placed in specific locations configured forefficient conversion of the absorbed optical energy into a short pulsesof acoustic energy within a wide band of ultrasonic frequencies; b)means for delivering said short ultrasonic pulses with known amplitudeand ultrasonic frequency spectrum through a coupling medium to a volumeof interest in a subject at a given time or time zero; c) means fordetecting said ultrasonic pulses in multiple positions at or around saidvolume of interest and measuring one or more parameters of time ofpropagation, amplitude and ultrasonic frequency spectrum, after saidultrasonic pulses are transmitted through or reflected from the volumeof interest using an array of wide-band ultrasonic transducers thatconvert ultrasonic pulses into electronic signals; d) means for analogamplification and digital recording of said electronic signals; e) meansfor performing signal processing to remove distortions of electronicsignals; f) means for image reconstruction using mathematical tomographyalgorithms; g) means for image processing and display; h) means for datatransmission and system control.

In this embodiment system may be configured to produce in real time at avideo rate two-dimensional images of thin tissue slices based onmeasured parameters of the speed of sound, ultrasound attenuation orultrasound backscattering. Also, in this embodiment system may beconfigured to produce three-dimensional images of the volume of interestin a subject body based on measured parameters of the speed of sound,ultrasound attenuation or ultrasound scattering. In an aspect of thisembodiment the means for detecting the ultrasonic pulses comprises ahand-held probe configured for acquisition, reconstruction and displayof real-time two-dimensional or three-dimensional images.

In another embodiment of the present invention there is provided adual-modality imaging system, comprising a) first means comprising thelaser ultrasonic system of described supra configured to generatetomographic images of a volume of interest in a subject body utilizingparameters comprising one or more of the speed of sound, ultrasoundattenuation or ultrasound backscattering; and b) second means forgenerating optoacoustic tomographic images of distribution of theoptical absorption coefficient in the subject body utilizing parametersof the absorbed optical energy density or various quantitativeparameters that can be derived from the optical absorption.

In this embodiment the first generating means may compriselaser-generated ultrasound and the second generating means may compriselaser-generated optoacoustics, both of said first and second meanscomprising an ultrawide-band ultrasonic transducer array positioned foracoustic detection of transient pressure waves resulting from deliveryof the laser-generated ultrasound and the laser-generated optoacoustics.Particularly, the images may be generated by the laser-generatedultrasound are tomographic images of tissue anatomy, morphology andstructure. In an aspect of this embodiment the images may be generatedby the laser-generated optoacoustics are tomographic images of tissuefunctional molecules such as hemoglobin, oxyhemoglobin, water, lipids,proteins and other molecules of biomedical interest. In another aspectthe images may be generated by the laser-generated optoacoustics aretomographic images of proteins, nucleic acids, enzymes and othermolecules comprising tissue of biomedical interest targeted withexogenous contrast agents or images of a spatial distribution of theexogenous contrast agents, where the contrast agents increasing contrastor characterizing molecules, cells or tissues. Representative examplesof exogeneous contrast agents are optical, optoacoustic, acousticultrasonic or dual optoacoustic-ultrasonic contrast agents and thecontrast agents are either molecules or nanoparticles. In allembodiments and aspects of the present invention the images may bespatially coregistered or temporally coregistered.

In yet another embodiment of the present invention there is provided aimaging method for increasing contrast, resolution and accuracy ofquantitative information obtained within a subject, comprising the stepsof a) producing a laser ultrasound or laser optoacoustic image of anoutline boundary of a volume of interest within the subject using thedual-modality imaging system described supra; b) generating a spatiallyor temporally coregistered image of speed of sound and/or an image ofultrasonic attenuation within the outlined volume boundary frominformation contained in the laser ultrasound or laser optoacousticimage; and c) generating a spatially or temporally coregisteredoptoacoustic image based on absorbed optical energy using an algorithmof the image reconstruction that employs distribution of the speed ofsound and/or ultrasound attenuation within the outlined volume boundary.

In yet another embodiment of the present invention there is provided alaser optoacoustic ultrasound imaging system (LOUIS), comprising a) adual laser source switchable between a laser ultrasonic mode and a laseroptoacoustic mode, said laser source capable to emit either shortoptical pulses with high repetition rate for the illumination of theultrasonic emitters in the ultrasonic mode or short optical pulses withlower repetition rate but higher pulse energy for the illumination ofthe volume of interest in the optoacoustic mode; b) an imaging modulecomprising one or more ultrawide-band ultrasonic transducers configuredto detect, through a coupling medium, optoacoustic and ultrasonicsignals propagated as transient pressure waves from said volume ofinterest within a subject body; c) means to rotate and/or translate saidimaging module relative to the volume of interest in the subject body tocreate multiple pressure waves, said means computer controllable ormanually controllable; d) means for processing said detected laseroptoacoustic and laser ultrasonic signals and reconstructing processedsignals into one or more of anatomical and functional/molecular imagesof the volume of interest in the subject body. The present invention isdirected to a related laser optoacoustic ultrasound imaging systemfurther comprising means for displaying the one or more images orsuperimposed coregistered images of the subject body or the volume ofinterest therein. Further to this embodiment the LOUIS imaging systemcomprises means for displaying the one or more images or superimposedcoregistered images of the subject body or the volume of interesttherein.

In both embodiments laser optoacoustic illumination may be performed inorthogonal mode, backward mode forward mode relative to the subject bodyor the volume of interest therein. Also, laser ultrasonication may beperformed in transmission or forward mode or in reflection or backwardmode relative to the subject body or the volume of interest therein orin a combination of the modes. In addition the laser wavelength may beabout 532 nm to about 1064 nm. Furthermore, the one or moreultrawide-band ultrasonic transducers may be configured to detectultrasonic signals with no or minimal reverberations. Further still thetransducer array may be interchangeable for acquisition of various typesof images in order to achieve greater contrast, resolution, orquantitative accuracy of either optoacoustic or ultrasonic images orboth.

Also, in both embodiments the means for processing and reconstructingsaid detected ultrasonic signals comprises one or more of electronicamplifiers with time-gain-control circuits; multichannelanalog-to-digital-converter with a field programmable gate array; andimaging module design and tomography algorithms configured toreconstruct quantitatively accurate volumetric images.

In one aspect of these embodiments the rotating means may be configuredto rotate the imaging module, wherein the detecting array of transducerscomprises an arc-shaped array or linear flat array or combination ofsaid array shapes comprising small ultrawide-band ultrasonic transducerswith wide angular directivity. In another aspect the translating meansmay be configured to translate said imaging module, wherein thedetecting array of transducers comprises an arc-shaped array or linearflat array or combination of said array shapes comprising finite sizeultrasonic transducers with narrow angular directivity. In addition, inthese embodiments and aspects, the imaging module comprises a hand-heldprobe configured for acquisition, reconstruction and display ofreal-time two-dimensional or three-dimensional images.

In yet another embodiment of the present invention there is provided amethod for imaging a subject's body or a volume of interest within,comprising the steps of a) positioning the subject body within orproximate to the imaging module of the laser optoacoustic ultrasoundimaging system described supra; b) delivering a laser-generated pulsesof ultrasonic energy to a volume of interest in the subject body; c)detecting the transmitted or reflected ultrasonic pressure waves whilemeasuring one or more parameters comprising a difference between thetime of emission and a time of arrival, a difference between emittedamplitude and detected amplitude, and a difference between ultrasonicfrequency spectrum of emitted and detected ultrasonic pulses; d)delivering a laser-generated pulse of optical energy to a volume ofinterest in the subject body; e) detecting the ultrasonic pressure wavesgenerated through optical absorption inside the subject body whilemeasuring one or more parameters comprising a time of arrival relativeto a time of generation, an amplitude of detected optoacoustic signals,and an ultrasonic frequency spectrum of detected optoacoustic signals;f) scanning the subject body or volume of interest therein with adetecting array of ultrawide-band ultrasonic transducers by repeatingsteps b) to e) at multiple positions around the subject body or volumeof interest while simultaneously scanning the sources of optical energyand sources of ultrasonic energy such that relative position of thedetecting array of ultrasonic transducers and the sources of optical orultrasonic energy can change or remain constant during the scans; g)processing the detected ultrasonic signals to remove distortions ofdetected signals; and h) reconstructing one or more volumetric imagesvia mathematical tomography algorithms using data of the processedsignals.

In this embodiment the pulse of optical energy may have a durationshorter than the time of pressure wave propagation through the distancein the subject body or volume thereof equal to a desired spatialresolution. Also, the other energy may be electromagnetic energy with awavelength of about 1 nm to about 1 m. In addition the one or morevolumetric images may be three-dimensional images of the volume ofinterest or of the subject body, or may be two-dimensional slicesthrough the three-dimensional volume of interest or even one-dimensionalprofiles of molecules of interest within the volume. Furthermore atleast one volume of interest may be a tumor, a lymph node, a vascularcirculation network, or a brain. Further still the laser ultrasound orlaser optoacoustic images may provide a feedback for guidance oftherapeutic treatments or surgical interventions.

In this embodiment the scanning step may comprise a) scanning the wholesubject subject body with a first array of ultrasonic transducers in arotational configuration to determine at least one volume-of-interestand its characteristics related to absorbed optical energy; b) replacingthe first array with a second array of ultrasonic transducers in atranslational configuration; and c) scanning through said at least onevolume-of-interest with a high resolution sufficient to acquirequantitative information related to distribution and concentration offunctional molecules therein. Also the step of delivering pulsed opticalenergy may be performed at multiple wavelengths of light, whether insequence or toggling.

Provided herein is a dual- or multi-modality three-dimensional (3D)tomography or imaging system that comprises laser optoacoustictomography (OAT) and laser ultrasound tomography (UST). Thisthree-dimensional tomography system provides comprehensive biomedicalinformation about a portion of the subject body under examination. Morespecifically, the system employs principles of laser ultrasound andlaser optoacoustic imaging to reconstruct three-dimensionaldistributions showing anatomical structures of a portion of the subjectbody under examination, molecular composition and distribution offunctionally important molecules in biological tissues of the subjectbody. All tomographic images are correlated and spatially coregistered.For dynamic processes that change over time, temporal coregistration canbe obtained so that anatomical and molecular images can be superimposedat a given time. Furthermore, optoacoustic images of the outline of thesubject body, i.e., the skin, are used to inform more accuratereconstruction of ultrasonic images, and the ultrasonic images in turn,inform more accurate reconstruction of optoacoustic images of thevolumetric distributions of molecules of interest.

The instant invention describes the full set of properties of the layersof the materials for the most effective generation of ultrawide-bandultrasound with laser pulses, not discussed in the prior art. They are avery small thickness of the layer of the laser illuminated materialmeasured in microns, a very strong optical absorption of a selectedlaser wavelength so that sufficient optical energy can be absorbed evenwithin the very small thickness of the layer, and a largethermo-acoustic efficiency parameter

${\Gamma = \frac{c_{0}^{2}\beta^{*}}{C_{p}}},$

for the material of the illuminated layer or large thermoacousticefficiency (often called Gruneisen parameter) of the medium surroundingthe laser-illuminated layer. The large Γβ can be achieved through alarge thermoelastic expansion coefficient, β, and fast (high) speed ofsound, and small heat capacity. These properties must be combined in onedesign to achieve maximum efficiency.

This invention provides a three dimensional tomography system thatacquires and displays comprehensive volumetric information aboutbiomedical object of interest, for example, tissue, cells, subject bodyor organ, with high contrast and high resolution. The depth at whichthis information can be obtained under optimal imaging conditions is upto 6-7 cm, which is significantly greater than the depth of pure opticalimaging with similar resolution. With this depth of imaging, biomedicalobjects such as human breast as large as 14 cm can be visualized. Theinformation that can be obtained from LOUIS images includes anatomical,i.e., structural or morphological, information and functionalinformation about hemoglobin distribution in blood and the level ofoxygenation in the hemoglobin. LOUIS also can provide images ofbiomedical objects with molecular specificity, i.e. images ofdistribution of molecules of interest.

If these molecules do not have sufficient intrinsic optical absorptionin the wavelength range of laser pulses utilized in LOUIS, then contrastagents targeted to those molecules through specific molecular probes orother high affinity vectors can be used. LOUIS contrast agents aremolecules, nanoparticles, nanobubbles or combination thereof. Theoptoacoustic ultrasonic contrast agents are in general those probes thathave high optical absorption and/or utilize high thermoacousticefficiency and/or have strong capability to scatter, reflect or absorbultrasonic waves or change speed of sound in the said biomedical objector any substance or structure that can be used to enhance contrast ofLOUIS images.

Ultrasound pulses for 3D biomedical imaging can be generated by shortlaser pulses, which gives significant advantages to the systemperformance and image contrast and resolution. Specifically, a specialultrasound generating medium, which under illumination of a short laserpulse produces clean smooth short non-reverberating pulses ofultrasound, is utilized. This produces either monopolar pressure pulses(so called Delta pulses of ultrasound (6)) or bipolar pressure pulses,if an application requires such pulses. Short nonreverberatingultrasound pulses produced by laser pulses or by pulses ofelectromagnetic energy, in general, will results in greater resolutionand contrast of 3D ultrasonic images. For example, a standardpiezoelectrically generated ultrasound pulse has 3-4 reverberations, soif produced with 12 MHz central frequency will have envelope frequency3-4 MHz effectively.

Therefore, the axial resolution of ultrasonic images is defined by thefrequency of an envelope of that reverberating ultrasonic pulse and beat least 3-4 times lower than that of 3D ultrasound image produced withlaser pulses. Short nanosecond laser pulses can generate pulses ofultrawide-band ultrasound with frequencies from low (tens of kHz) tohigh (tens of MHz). These ultrawide-band ultrasound pulses are verybeneficial for ultrasound imaging since they is effectively scatteredand attenuated by variety of biomedical object structures (large such astumors or large vessels to small such as microvessels to microscopicsuch as cells and even subcellular components. Biomedical objects(tissue and cells) can absorb and scatter certain frequencies ofultrasound while other frequencies can pass said objects undistorted.Therefore, spectroscopic analysis of laser ultrasonic signals in termsof their frequency spectra can reveal useful diagnostic information.Three dimensional images obtained with laser ultrasound such as theimage ultrasound attenuation, the image of ultrasoundscattering/deflection and the image of distribution of ultrasoundvelocity (most frequently called speed of sound) are also very rich ofinformation that can be used by physicians and biomedical researchersfor characterization and differentiation of biomedical objects (tissues,cells, organs etc).

LOUIS utilizes short nanosecond laser pulses for generation of shortpressure pulses which propagate as ultrawide-band ultrasound inbiomedical objects. LOUIS operates in two modes, Laser Ultrasonic andLaser Optoacoustic. Images of both modes can be fully coregistered,correlated and superimposed since they are collected with one and thesame set or array of ultrasonic transducer detectors. In general, LOUIScan utilize illumination with any optical wavelength or even anywavelength of electromagnetic energy and any sequence or duration ofpulses of said electromagnetic energy. But short, about 1 ns to about 20ns laser pulses in the near-infrared spectral ranging from about 650 nmto about 1250 nm are preferred for imaging with LOUIS.

In the laser ultrasonic mode, the laser pulses illuminate a specialmedium placed outside of the biomedical object of interest, so thatthese short pulses of ultrasound enter the biomedical object ofinterest, propagate through the object of interest and interact with theultrasonic transducers for purposes of their detection. A laserwavelength selected for generation of laser ultrasound pulses is usuallychosen to be strongly absorbed in the external special medium and theneffectively converted into heat and pressure, with high-pressuregeneration efficiency being the ultimate goal. The detected ultrasonicpulses represent electronic signals that, after signal processing, e.g.,filtering, conditioning, analysis etc., are used for furtherreconstruction of volumetric ultrasonic images using mathematicalalgorithms. LOUIS can be used to reconstruct at least three types ofultrasonic images: the image of the speed of sound, the image ofultrasonic attenuation and the image of ultrasonic reflection(deflection, scattering).

In the laser optoacoustic mode the laser pulses illuminate thebiomedical object of interest itself, propagate through the object andinteract with the object of interest, so that the energy of theseoptical pulses can be absorbed by its components and constituents andconverted into heat and simultaneously thermal pressure, which thenpropagates as ultrasound and interacts with said ultrasonic transducersfor purposes of their detection. The wavelength of the laser pulses isselected to propagate to a desirable depth in the object, e.g., tissue,and become preferentially absorbed by specific molecular constituents ofinterest: hemoglobin, oxyhemoglobin, water, lipids, melanin and otherendogenous molecules of interest or exogenous molecules or particles orprobes of exogenous contrast agent.

The detected ultrawide-band ultrasonic pulses represent electronicsignals, which after signal processing, i.e., analysis, filtering,conditioning, etc, are used for further reconstruction of volumetricoptoacoustic images using mathematical algorithms. The optoacousticimages represent distribution of absorbed optical energy at a selectedwavelength or a collection of multiple wavelengths, and afternormalization to distribution of the optical fluence can representdistribution of the optical absorption coefficient in the biomedicalobject. After image post-processing the optoacoustic images can beconverted into a number of quantitative volumetric images, including,but not limited to. the following five types: the image of the totalhemoglobin (THb), the image of hemoglobin oxygenation (SO2), the imageof water distribution (H2O), and the image lipid/fat distribution(Lipid) and the molecular image of distribution of a specific moleculeof interest.

In order to transmit ultrasonic and laser (optical) pulses to thebiomedical object, then detect ultrasonic (acoustic pressure) pulsesfrom the object and reconstruct the laser ultrasound and laseroptoacoustic images using LOUIS, usually a coupling medium is required.For better image quality the following properties of the coupling mediumis desired: good optical transparency in the wavelength range of laserpulses used for illumination, good ultrasonic acoustic transparency inthe frequency range of ultrawide-band ultrasonic pulses used forimaging, good matching of the optical refraction index to the tissue ofthe biomedical object and good acoustic impedance matching to the tissueof said biomedical object. In addition, it will help to image deeper andwith less noise and artifacts, if the coupling agent makes the tissue ofthe biomedical object optically clear. Skin clearing media have beenproposed and developed for increased optical transparency of skin forbetter quality of optical images. However, as disclosed herein, opticalclearing agents can improve quality, fidelity and contrast of laseroptoacoustic images and laser ultrasonic images.

Many types of lasers and other pulsed sources of electromagnetic energycan be used for LOUIS. The most preferred lasers are those tunable inthe near-infrared spectral range and simultaneously robust forbiomedical applications, such as Nd:YAG pumped Ti:Sapphire laser andsolid state diode laser matrices.

The ultrasonic transducers (detectors) can be made of various materialsand utilize various technologies. The preferred materials includepolymers, crystals, ceramics, and composites. The types of ultrasound(pressure) detectors include piezoelectric transducers, capacitivemicromachined ultrasonic transducers (CMUT), optical beam deflectiontransducers, fiberoptic sensors, optical interferometers andmicrophones. The most preferred detectors for LOUIS are those thatpossess higher sensitivity and simultaneously can detect ultrasoundwithin an ultrawide band of ultrasonic frequencies.

Signal processing in LOUIS includes analysis of signal profiles, signalamplitudes and spectrum of signal frequencies. Spectra, e.g., Fourierspectra, of laser ultrasound signals propagated through the biomedicalobject can be analyzed to reveal properties of tissues important forbiomedical diagnostics. Such spectra of laser optoacoustic signalsgenerated by optically induced acoustic sources within the biomedicalobject and propagated through the biomedical object also can be analyzedto reveal properties of tissues important for biomedical diagnostics.

Analysis of noise in the system can help to filter the noise and improvecontrast of images. Whether the noise is white and noncorrelated or thenoise is correlated between various detectors or transducers ortransducer positions around the object, mathematical methods exist andcan be chosen to provide the best filtering of the signals from noise.In general, signal processing for LOUIS is designed to reverse the socalled system transfer function, i.e. all distortions that introducedinto the detected ultrasonic signals by the system components, such aslasers, detectors and analog and digital electronics. The goal is toobtain electronic signals with properties as close as possible to theintrinsic pressure or ultrasound signals.

One specific method of signal processing is preferred due to theaccuracy of quantitative information provided by the volumetricoptoacoustic images. This method provides for volumetric imagereconstruction based on signal deconvolution using the Curvelettransform, a two-dimensional wavelet transform, known in the art, forfiltering optoacoustic and ultrasonic signals. The most desirableproperty of wavelets is their capability to filter signalssimultaneously in time and frequency domains, thus providing greatseparation of useful signals and noise that appear in the same frequencyrange. Thus provided herein is an algorithm for laser ultrasonic andlaser optoacoustic image reconstruction in 3D using the Curveletdeconvolution method. Also provided are algorithms aimed at totalvariance minimization that can be beneficial for laser ultrasound andlaser optoacoustic tomography.

Three dimensional tomography images are much more quantitativelyaccurate compared with two-dimensional images due to collection ofcomplete sets of data and to rigorous reconstruction algorithms based oninformation about the object collected from various angles and positionsin the 3D space. The ultimate image would be a 3D image obtained in realtime, i.e. obtained within such a short period of time when importantbiomedical conditions of the object of interest could not change.Typically, acquisition of 10-30 images per second in biomedicalapplications is sufficient to be considered as real-time monitoring. Oneimage per second also is acceptable for monitoring kinetics and dynamicsof biological processes. So, the most important are designs in whichdata are collected rapidly, while image reconstruction can be donelater. Alternatively, image reconstruction in real time brings practicalconvenience in biomedical imaging, allowing the doctor to make animmediate decision in the presence of a patient. Thus, the presentinvention provides reconstruction of laser ultrasonic and laseroptoacoustic images with hardware and algorithms operating in real timewith the use of the modern and advanced computer power capabilities.Field Programmable Gate Arrays (FPGA) microprocessors are most effectivefor signal processing, Graphical (multicore) processor units (GPU) aremost effective for image reconstruction, while the Central ProcessingUnit (CPU) of a computer is the most effective for display of images andsystem controls.

Thus, LOUIS has multiple biomedical applications including but notlimited to, cancer detection or screening, including detection of cancerin the lymph nodes and metastatic tumors, cancer diagnostics, monitoringeffects of anticancer therapy and aggressiveness of a cancer, detectionand characterization of vascular diseases, such as, cardiovasculardisease, stroke, peripheral vascular disease, diseases that result inthe damage of microvasculature, e.g., diabetes, atherosclerosis,monitoring circulation and its functions, anatomical, functional andmolecular characterization of various tissues and health conditions,functional imaging of blood distribution and its oxygen saturationlevels. Other biomedical applications include molecular imaging ofvarious molecular targets of diseases and otherwise abnormal tissues,monitoring kinetics of drug distributions and biodistribution ofnanoparticles and other contrast agents, monitoring physiological andpathological processes in the animal or human subject body, monitoringtrauma, burns and otherwise damaged tissues and the process of itsrecovery after treatment.

Particularly, the combined imaging system comprises the followingadvantages:

LOUIS—Combined 3D Optoacoustic/Ultrasonic Imager

Laser optoacoustic ultrasonic imaging system is a 3D tomography systemfor the comprehensive characterization of biomedical objects. The 3Dtomography system creates a spherical surface of virtual transducers byrotation of an arc-shaped ultrasonic array around the object of interestwith computer-controlled illumination from multiple positions, whichpermits the most beneficial distribution of light in the object. Thetime of the entire 3D image acquisition can be as short a few seconds,but may be extended for several minutes for the benefit of image qualityin the object has low contrast. The LOUIS system components compriseelectronics hardware, firmware, software and custom designed wavelengthtunable lasers. One laser has relatively low pulse energy of about 0.1to about 2 mJ, and a high repetition rate of laser pulses (1-5 kHz) usedto generate ultrasound pulses outside the subject body underexamination. The second laser has much higher pulse energy, up to 250mJ, a relatively low repetition rate (10-20 Hz) and a wavelength tunablein the near-infrared spectral rage, with capability to electronicallyswitch or toggle the illumination wavelengths, for example, 1064/800 nm,1064/757 nm, for functional optoacoustic imaging.

Use of Laser-Induced Ultrasound for UST

Conventional electrical generation of ultrasound was replaced withlaser-induced ultrasound (LU) for transmitting short ultrasound pulsesto the breast and thereby achieving three-fold improved UST imageresolution and greater sensitivity. LU is emitted by a thin layer ofblack polydimethylsiloxane (PDMS) or, alternatively, polymethylmethacrylate (PMMA) filled with absorbers polymer embedded with highlyconcentrated absorbers. Strong absorbers are, but not limited to, carbonnanotubes, strongly absorbing in the near-infrared and having highthermal expansion coefficient. This thin layer is illuminated by pencilbeams of short (8 ns) laser pulses from Nd:YAG laser. To decrease thedata acquisition time for laser ultrasound imaging, a diode laser can beused with pulse repetition rate of about 1-5 KHz, pulse energy of about1-2 mJ and pulse duration of 1-3 ns. As a result of strong opticalabsorption thermal pressure is generated by point sources resulting inspherical ultrasonic waves with ultrawide bandwidth from about 50 KHz toabout 30 MHz. The first application of LU was performed in phantoms toobtain fully 3D UST images.

Novel Optoacoustic/Ultrasonic Transducer Array as LOUIS Imaging Probe

Current commercial medical ultrasonic transducers provide spatialresolution two-three times lower than potentially attainable with agiven ultrasound frequency. The invented new technology of ultra-wideband transducers we teach here improves sensitivity to enable theoptoacoustic imaging of tumors at significant depth up to 6-7 cm, i.e.through large biomedical objects such as entire breast, and alsoimproves resolution of ultrasound images. With novel transducermaterials employed in our probes we achieved a very challenging goal:increase sensitivity of detection and simultaneously increase thedetection bandwidth.

Advanced 3D Image Reconstruction Methods

New image reconstruction algorithms are developed and implemented forforming images that depict the distribution of the absorbed opticalenergy density within biomedical objects (live tissues), which canreveal the location of cancerous lesions or other abnormalities thathave elevated blood content. Both analytic and iterative reconstructionalgorithms are developed and quantitatively evaluated for performance.These algorithms compensate for important physical factors such as theimpulse response of the transducer, stochastic and acoustic noise, andfinite sampling effects.

I. Dual Mode Image Reconstruction and Coregistration of 3D UST with 3DOAT

In addition to recording optoacoustic signals for use in OAT, thedeveloped 3D imager (LOUIS) is capable of operating in 3D laser USTmode. This enables a novel 3-step method for image reconstruction andprocessing, which results in significantly higher contrast andresolution of coregistered images. At the first step, we acquire data isacquired and an optoacoustic image or ultrasonic image of the outline ofthe subject body part under examination is reconstructed. This permitsaccurate separation of the two domains: subject body part underexamination and surrounding volume of the coupling agent. At the secondstep, data are acquired and image reconstruction methods are implementedfor forming images that depict the 3D speed-of-sound (SOS), attenuation,and reflectivity distributions in the portion of the subject body underexamination, outlined and defined on the image obtained in the firststep.

Therefore, the image of the first step informs a more accuratereconstruction of the image obtained in the second step. At the thirdstep, a volumetric optoacoustic image of the subject body underexamination is acquired and reconstructed using information contained inthe image obtained in step 2. For example, an image of the speed ofsound distribution can be used to correct the time of arrival ofoptoacoustic signals and thus reconstruct more accurate optoacousticimages. In general, the image providing anatomical/structuralinformation can inform more accurate reconstruction of optoacoustic orfunctional images. The two types of images (anatomical and functional)are complementary. This is achieved by developing specialized imagereconstruction algorithms that utilize boundary conditions andregularization constrains determined from images reconstructed in theprevious step.

Preferably, the combined imaging system comprises the physicalstructure, methods utilized during imaging and the hardware, softwareand algorithms described below.

Dual-Modality Laser Optoacoustic/Ultrasonic 3D Tomography Imager

The design of the imaging module (see FIGS. 3A-3B) and its componentsimproves, extends and significantly enhances of previously developedpreclinical 3D OAT imager (21). The imaging module provided herein,contains a 128 element ultrasound detector array and 7 optical fiberbundles, 4 of which are used for optical illumination of the biomedicalobject inside the module and its optoacoustic imaging, and 3 of whichare coated with a thin absorbing polymer layer to generate laserultrasound and acquire different types of ultrasound images (speed ofsound, ultrasonic attenuation and ultrasound scattering. This uniquedesign enables three different types of measurements to be acquiredduring a single imaging study: 1) optoacoustic signals for OAT imagereconstruction at different laser wavelengths; 2) deflected orbackscattered ultrasound for reconstruction of ultrasonic reflectivitymaps; and 3) transmission ultrasound for reconstruction of ultrasonicSOS and attenuation maps. The entire imaging module will rotate in orderto collect tomographic measurements that are sufficient for accurateimage reconstruction.

The ultrasound array is arc-shaped with radius of 70 mm and angularaperture of 150 deg. The remaining 30 deg opening is used for suspendingthe biomedical object, such as a breast in prone downward position or awhole small animal. The probe has 128 transducers with lateraldimensions of 1.3 mm×1.3 mm and a pitch of 1.4 mm. The transducers aresensitive within an ultrawide band of ultrasonic frequencies from 100KHz to 10 MHz and exceptionally sensitive in allowing detection of 1 Papressure with signal-to-noise (SNR) of 2.

Another novel component of the imaging system is the use oflaser-produced ultrasound (LU) for insonifying the breast, as opposed totraditional electrically produced ultrasound (31). LU is emitted by athin layer of PMMA polymer with embedded highly concentrated absorbers,for example, carbon nanotubes, strongly absorbing in the near-infraredand having high thermal expansion coefficient. This thin layer isilluminated by pencil beams of short (8 ns) laser pulses from Nd:YAGlaser. As a result of strong optical absorption, thermal pressure isgenerated by point sources resulting in spherical ultrasonic waves withultrawide bandwidth from ˜50 KHz to about 30 MHz. The ultrasonic pulsereplicates the shape of the laser pulse, which is smooth and short andhas no reverberations typical of electrically generated ultrasound. Ofcourse, very high frequencies above 12 MHz can be lost in propagationthrough tissues, but 12 MHz pulse without reverberations will produceultrasound resolution equivalent of reverberating 30-35 MHz pulses.

There are three main advantages of employing Laser Ultrasound (LU) asopposed to electrically (transducer) produced in the dual- ormulti-modality imager: 1) better spatial resolution, 2) bettercontrast/sensitivity, 3) simpler and low noise electronics, that is notransmit/receive switches. Image spatial resolution can be superiorbecause LU produces clean, smooth short pulses of ultrasound, not thetypical reverberating pulses of electrically generated ultrasound, whichneeds to be enveloped for imaging purposes. Image contrast can beenhanced because LU pulses have relatively high intensities and minimumbackground noise. The system electronics are simplified because they areonly used for read-out. This circumvents the need to emit 200 V pulsesand then quickly detect microVolt signals. Transmit/receive switches arethe main source of noise in the conventional ultrasound systems. Forexample, a supersensitive amplifier sitting next to a super powerfulemitter-amplifier can easily be saturated with noise.

Provided herein are examples of LOUIS images of a whole mouse subjectbody (see FIGS. 11A-11B). It was demonstrated previously that softtissue organs, spine, ribs and joints, vasculature or microvasculaturecan be clearly visualized (21). Microvasculature as small as 50 micronwas visualized, even though spatial resolution of the instant system isabout an order of magnitude lower.

Thus, the present invention demonstrates the feasibility of a 3Dtomographic system design for performing dual-mode laser optoacousticand laser ultrasonic tomography. LOUIS tomography system creates aspherical surface of virtual transducers by rotation of an arc-shapedultrasonic array around the object of biomedical interest withcomputer-controlled illumination from multiple positions, which permitsthe most beneficial distribution of light in the object. For performingultrasound tomography, conventional electrical generation of ultrasoundfor object insonification is replaced with laser-produced ultrasound,thereby, resulting in a three-fold improvement in image resolution. Thesystem development includes electronics hardware, firmware, software andcustom design a multi-wavelength tunable laser that enables thecapability to electronically switch or toggle the illumination colors,e.g., 1064 nm and one NIR wavelengths in the range from 730 to 850 nm,for optoacoustic imaging. This permits differential imaging of variouschromophores, such as hypoxic and oxygenated blood.

OAT image reconstruction algorithms are implemented in LOUIS for formingimages that depict the distribution of the absorbed optical energydensity within biomedical object, which can reveal the location ofabnormal tissues such as cancerous lesions that have elevated bloodcontent. Both analytic and iterative reconstruction algorithms aredeveloped and quantitatively evaluated (see below detailed descriptionof math physics algorithms). These algorithms compensate for importantphysical factors such as the response of the transducer, stochastic andacoustic noise, and finite sampling effects.

Laser ultrasound tomography utilizes our image reconstruction methodsfor forming images that depict the 3D speed-of-sound (SOS), ultrasoundattenuation coefficient, and reflectivity distributions of biomedicalobject or organ tissue. These images provide structural information thatis complementary to the functional (blood content and oxygenation)information conveyed by the OAT images. Moreover, we teach that thereconstructed SOS and attenuation maps can be utilized to furtherimprove the accuracy of the reconstructed OAT images. This can beachieved through specialized OAT reconstruction algorithms thatcompensate for variations in the SOS and attenuation distributions.

Computer Modeling

Imager development is based on a comprehensive computer model of 3D OATand UST. This model includes the following components: 1) calculation ofthe distribution of absorbed optical energy exponentially decreasing indepth of the breast (32-34), 2) generation of optoacoustic signals, 3)generation of LU for UST imaging, and 4) calculation of profiles ofdetected signals taking into account the geometry of each transducerelement, i.e., directivity diagram of each element, and sensitivity ofpiezoelectric detectors as a function of the ultrasonic frequency, i.e.,effect of bandwidth (29). Computer-software has been developedpreviously by the inventors that is utilized for establishing acomprehensive physics-based model of the imager. The hardware design isconducted concurrently with the designs of the image reconstructionalgorithms described below, so that they can be informed and refinedjointly. The image quality measures used to guide the system refinementsare described below.

I. OAT Detection-Sensitivity

The sensitivity of optoacoustic detection depends on the product of 4parameters: the effective optical fluence acting on the tumor, theoptical absorption coefficient of the tumor, the thermoacousticefficiency F, i.e. the ability of tissue to convert light intoultrasound, and the sensitivity of the piezoelectric transducer (35).Using the experimentally measured sensitivity of our new transducers,about 15 microVolt/Pa, and optical properties of breast tumors andnormal tissue previously obtained, one can calculate minimal detectableblood content in a tumor with defined dimensions and depth from theilluminated surface (36). Based on this calculation, the imager iscapable of detecting not only tumors with dimensions of about 10 mmregularly found by mammography screening, but also early tumors having avery small size of 3 mm. While the detection sensitivity will degradewith depth, those very small tumors may be detected at a depth of 6-7 cmdepending on the density of tumor angiogenesis, which in turn definesoptical absorption of the tumors (37-39).

II. OAT Imaging Depth

The anticipated imaging depth of OAT in the breast is about 6 cm fortypical 10 mm tumors and about 8 cm for blood vessels dependent on theHb concentration and their dimensions, i.e. comparable with the imagingdepth of high-resolution (12 MHz) breast B-mode ultrasound. Even thoughbreast tumors statistically occur most frequently at the depth of 1-3cm, herein the maximum depth of detection is about 6 cm due toinfrequent occurrence of deep tumors in very large breasts. Havingeffective optical attenuation in tissue of about 3 times per cm ofdepth, the optical fluence is attenuated about 729 times before it canreach 6 cm depth. However, system electronics described herein isdesigned with a dynamic range of 14 bits, which permits simultaneousdetection of maximum signals and signal attenuated more than 4 orders ofmagnitude. Furthermore, ultrasensitive transducers provided herein candetect pressure levels of about 1 Pa with signal-to-noise ratio of about2 (40,41). A 2 Pa pressure can be detected from a ˜1-mm object, e.g. ablood vessel, with the optical absorption coefficient of 10/cm locatedat the depth of 8 cm from the breast tissue surface illuminated with anear-infrared laser pulse having safe optical fluence of 20 mJ/cm² (8).

III. Spatial Resolution for OAT and UST

Previously, microvessels as small as 50 micron were visualized in apreliminary design of LOUIS animal imager (21), even though spatialresolution of that system was about an order of magnitude lower. Thespatial resolution of the OAT images can be spatially variant, beingworse at locations that are near the measurement transducer (6,23). Theworst spatial resolution for the OAT image, as measured by the FWHM of apoint-source response (42), is 0.5 mm. The resolution of thereflectivity UST image is limited by half the effective wavelength,which results in spatial resolution significantly better than 0.5 mm.The resolution of the SOS and attenuation UST images is largely limitedby the density of transmit-receive pairs (i.e., number of tomographicviews) and the efficacy of the image reconstruction algorithms. Anapproximately isotropic spatial resolution of <1 mm is presentlydemonstrated for LU part of LOUIS (see FIG. 8B).

IV. UST Reconstruction Accuracy

Using well-calibrated phantoms enabled reconstruction of the ultrasonicSOS and attenuation distributions of subcutaneous fat, glandular tissue,and tumor tissue to within 0.2% of their known values. Similartolerances have been reported in studies of breast UST (13,17,18). Theultrasonic reflectivity image is typically used to reveal tissueinterfaces only. LOUIS is capable of detecting not only boundaries butalso volumes of structures within the biomedical objects.

V. Data-Acquisition and Image Reconstruction Speeds

The acquisition speed for a full set data with a rotating arc-shapedprobe is about 3 minutes and multi-modality data acquisition time toless than a minute with the increased sensitivity of the noveltransducers and reduced number of averaged signals. The time for full 3DOAT image reconstruction, using a filtered backprojection algorithm,with resolution of 500 micron is reduced in the present LOUIS softwarerelative to earlier version to about 15-30 sec, depending on the totalnumber of voxels within the reconstructed volume, with application ofthe reconstruction software based on CUDA code and multi-core graphicsprocessing units (GPUs). Fully 3D image reconstruction of the UST imagesis accelerated using GPUs with an initial accomplishment ofreconstructing images in <10 min.

LOUIS Imager Hardware I. Transducer Array

The optoacoustic/ultrasonic transducer array provided herein is theprimary basis for novelty of the LOUIS imaging module. This criticallyimportant system component for hybrid dual-modality imaging must satisfya number of requirements. The optoacoustic signals contain acousticfrequencies ranging from about 200 kHz to 12 MHz, depending ondimensions of tissue optical heterogeneities in the breast and the laserpulse duration (40). Such ultrasonic waves propagate in tissues withattenuation that may be accounted for and deliver spatially resolvedinformation to the surface of tissue where they can be detected and usedfor image reconstruction (9). However, undistorted detection ofultrasound comprising such a wide frequency range requires acoustictransducers with an exceptionally wide bandwidth (43-45).

Ideally, an optoacoustic transducer is sensitive to the entire range ofacoustic frequencies to detect the small and large tissue structureswith resolution of <0.5 mm sufficient for biomedical imagingapplications in the depth of tissue. Therefore, new piezoelectricmaterials are incorporated into the design of the transducer arrays thatis part of a specially developed clinical probe. The composition of thepiezoelectric material and the design of a matching front layer andbacking material plays a major role in determining the bandwidth of theprobe. Extensive preliminary tests were performed with two differentpiezoelectric materials: single crystal PZT ceramics, leadmetaniobate-titanate (PMN-PT), modified lead titanate (MPT) as part of1-3 composites. Results demonstrated significant widening of thebandwidth and absence of reverberations in the novel transducersrelative to commercial ultrasonic transducers.

II. Patient Bed and Imaging Module for Breast Imaging

A patient table is constructed that contains an imaging module mountedunderneath. The patient lies in the prone position on the examinationtable with the breast suspended through an opening into an imagingmodule. In order to minimize motions of the breast in the imagingmodule, we design an inflatable ring balloon that shapes the breastcloser to its natural spherical shape. The height of the table isapproximately 45 in, which allows the system operator to visuallyposition the breast within the imaging probe while being seated. Nocompression is required and the breast is centered in the center of theimaging tank by minor movement of the patient. The imaging tank isfilled with warm clean water and appropriate plumbing is included in thedesign to permit rapid changing of the tank water.

III. Electronics, Firmware, Rotation Stage

The system electronics and computer controls in LOUIS are all upgradedfrom a prior OAT imager to minimize electronic noises. The rotationstage mechanism is essentially different to the one used in ourpreclinical imager and it provides more accuracy and capability toperform series of scans in clockwise and anticlockwise directionswithout loss of home position.

Methods I. Robust OAT Image Reconstruction Methods

In this component of LOUIS, OAT image reconstruction methods aredeveloped, implemented, and optimized for forming images that depict the3D distribution of the absorbed optical energy density within breasttissue by use of measurement data recorded by the imager. Two classes ofreconstruction methods are developed that permit different trade-offsbetween data-acquisition time and image reconstruction time.

II. Data-Restoration Methods for Use with Analytic ReconstructionMethods

Analytic OAT reconstruction algorithms, such as filtered backprojection(FBP) algorithms (46), form an image by numerically computing aclosed-form mathematical formula. Such methods can be computationallyefficient and yield relatively short accelerated reconstruction times,for example, <1 min for a volumetric image. However, they typicallyrequire a densely sampled tomographic data set to be acquired, which canextend data-acquisition times. Another shortcoming is that they arebased on idealized models that do not compensate for noise, theinstrument response, and other complicating factors related to theimaging physics.

The effectiveness of the computationally efficient 3D FBP algorithms wasimproved by developing novel methods for pre-processing the measuredmulti-dimensional optoacoustic signals prior to image reconstruction.This process is analogous to what is called “sinogram restoration” inthe X-ray CT community. This method has never been utilized foroptoacoustic and laser ultrasonic imaging. Specifically, robust methodsinspired by compressive sampling theory are developed to compensate forthe effects of the transducer impulse response and thermoelectricalnoise in the measured data. Methods for estimating missing data also aredeveloped, which require less data to be acquired and result inshortened the total imaging times. After the measured data have beenpre-processed, a computationally efficient 3D FBP algorithm is employedfor quantitative image reconstruction. The methods that are employed forachieving this are summarized below.

III. Sparsity-Regularized Data Restoration

An ultrasonic transducer's electromechanical impulse response (EIR)describes how its electro-acoustical properties degrade the recordedpressure data (47). In order to reconstruct an image that accuratelydepicts the absorbed optical energy density in OAT, the effect of theEIR on the measured optoacoustic signals must be accounted for. A robustmethod for measurement denoising and deconvolution of the EIR in OAT hasbeen designed. This method decovolves the EIR by solving the followingconstrained optimization problem:

{circumflex over (α)}=argmin∥α∥₁subject to ∥p−HC ⁻¹α∥₂≤ε  (1).

Here, α is the vector of expansion coefficients that correspond to thepressure data p=C⁻¹α, C⁻¹ is the synthesis operator that relates the 3Dpressure signal (two spatial coordinates plus time) to the expansioncoefficients, and H is an operator that describes a 1D temporal blurringof the pressure data due to the EIR. The parameter ε describes the noiselevel in the measured optoacoustic signals. The final estimate of thedeconvolved pressure data is obtained as {circumflex over(p)}=C⁻¹{circumflex over (α)}. The expansion functions used to representthe pressure data, which determines the explicit form for the operatorC⁻¹, are chosen such that expansion coefficient vector α=Cp is sparse.Such expansion functions include curvelets. Although curvelet transformis known from the prior art (48), the method has not been developedbefore now for optoacoustic image reconstruction. Efficient andnumerically robust algorithmic realizations of Eq. (1) are hereindeveloped and optimized. Methods for estimating missing measurements ofthe pressure wavefield is developed by use of a generalization of Eq.(1) that has proven effective for a similar application in geophysicalimaging (49).

This represents a fundamentally different approach for deconvolving theEIR in OAT. Specifically, the method is distinct from existing methodsused in OAT in that it exploits sparsity of the pressure data in asuitably defined transform domain, and exploits the fact that thepressure signal produced by an optical absorber will yield a continuouswavefront in the measured data space. Similar methods have been employedfor processing geophysical data (49) with great success. Results of thismethod using our new LOUIS imager are displayed in FIG. 19B. Use of theproposed method resulted in dramatically improved visibility of theblood-filled vessels and positive-valued pixel values that wereproportional to the absorbed optical energy density within the tissue.

IV. 3D Iterative OAT Reconstruction Methods

The data restoration methods discussed supra facilitate accurateanalytic image reconstruction. However, it is contemplated thatiterative OAT reconstruction algorithms can improve diagnostic imagequality for breast imaging applications. Iterative reconstructionalgorithms offer the possibility to compensate for noise, instrumentresponse, and other complicating factors related to the imaging physics.Iterative algorithms can mitigate data incompleteness, therebypermitting reduced data-acquisition times, but are more computationallyburdensome than analytic methods, such as the FBP algorithm. An FBPalgorithm is utilized to reconstruct an initial image for rapid viewing,while an iterative algorithm is utilized to reconstruct an improve imageoff-line for viewing at a later time (see FIGS. 20A-20B).

V. Limited Data Image Reconstruction

Iterative image reconstruction methods were developed based onconstrained total-variation (TV) minimization (50). The idea ofconstrained TV-minimization has proven useful in the field ofcompressive sensing, and is effective when there exists some sparserepresentation of the object. Iterative reconstruction algorithms fortomography that operate via L₁-norm minimization of the total variation(TV) of the object, subject to data consistency and object positivityconstraints were examined. These results suggest that for certainclasses of objects our reconstruction algorithms based onTV-minimization can significantly outperform conventional iterativealgorithms, yielding informative images even when the measured data arehighly incomplete. Other image reconstruction methods (51) inspired bycompressive sampling are also adapted and explored for 3D OAT asdescribed below. The developed algorithms compensate for the transducerEIR and also for the finite detection area of the transducer. Theinventors have developed a methodology for modeling the response of anultrasound transducer in iterative image reconstruction (29).

VI. Implementation on Graphics Processing Units (GPUs)

Because fully 3D iterative OAT image reconstruction can becomputationally demanding, it is necessary to implement the developedalgorithms using GPUs. Our team has specific expertise in theimplementation of OAT image reconstruction algorithms using the NVidiaCUDA programming environment. To demonstrate the speed-up factors thatcan be obtained, a preliminary study was conducted using an 8-core IntelXeon processor workstation clocked at 2.40 GHz equipped with 48G memoryand one NVIDIA Tesla C2050 GPU card with compute capability 2.0. An OATexperiment was simulated in which 360 transducers were evenlydistributed on a measurement circle with 20 cm radius, and eachtransducer collected 256 samples at 2 MHz sampling rate. A 2D numericalphantom (256×256) was employed to represent the optical absorptiondistribution. Image reconstruction was performed by minimizing aleast-squares cost function using a conjugate gradient method. The runtime of the GPU code was 30 seconds while our CPU code took 1755 secondsto complete the reconstruction, resulting in a speed-up factor ofapproximately 60 for the GPU-based code. The cross-correlation of thetwo images was computed to be 0.9997, indicating that there was not asignificant loss of accuracy by use of the GPU-based code. Ourexperience in this area permitted us to develop computationally feasible3D reconstruction algorithms that facilitate their clinicalapplications.

VII. 3D UST and UST-Guided OAT Image Reconstruction Methods

3D UST image reconstruction methods are established for use with thedeveloped multi-modal imager. Specialized UST-guided OAT reconstructionalgorithms that compensate for variations in the SOS and attenuationproperties of breast tissues were developed and implemented.

VIII. Reconstruction Methods for Sparse-Array 3D Ultrasound Tomography

Reconstruction methods are developed to form accurate images of the 3Dacoustic properties of the breast. As described below, methods aredeveloped for reconstructing images of three complementary breastproperties: SOS, acoustic attenuation, and reflectivity. These 3D imagesprovide a comprehensive description of breast anatomy that iscomplementary to the functional information revealed by the OAT image.These reconstruction methods account for problems that includemitigation of data incompleteness and noise and computationallytractably modeling of the relevant wave physics.

A. Reconstruction of SOS Distribution

Algorithms are developed for reconstructing the 3D SOS distribution thebreast from knowledge of time-of-flight (TOF) measurements of thetransmission ultrasound signals. Geometrical acoustic-based ray theoryis utilized to establish a non-linear model that relates the measuredTOF values to 3D SOS distribution as

$\begin{matrix}{{{{TOF}\left( {r_{s},r_{d}} \right)} = {\int_{L}{\frac{1}{c(r)}{dr}}}},} & (2)\end{matrix}$

where TOF(r_(s),r_(d)) is the TOF measured between source location r_(s)and detector position r_(d), c(r) is the sought after SOS distribution,and L=L(r_(s),r_(d);c(r)) is the curved path traveled by the acousticwave (that also depends on c(r)). For a given c(r), the Eikonal equation(52) is solved numerically to determine the ray path L. An iterativereconstruction method is developed for inverting Eq. (2) thatalternatively updates the estimates of c(r) and L and minimizes aregularized cost function to obtain the final estimate of c(r) It iscontemplated that further development of algorithms can be guided bybent-ray ultrasound tomography that has shown promise in pre-clinicalstudies (16,17).

B. Reconstruction of Attenuation Distribution

Algorithms are developed for reconstructing the 3D acoustic attenuationdistribution of the breast from transmission measurements. Accuratereconstruction of the acoustic attenuation requires knowledge of the SOSmap and is therefore be conducted after the SOS map is determined usingthe methods described above. Given that the SOS map is known, a linearimaging model is obtained as

a(r _(s) ,r _(d))=∫_(L)α₀(r)dr  (3),

where L=L(r_(s),r_(d);c(r)) denote the same ray paths as determined fromthe last iteration of the alternating SOS reconstruction describedabove, and α₀(r) is a acoustic attenuation coefficient (31). The datafunction α(r_(s),r_(d)) is determined as an energy ratio between themeasured transmission acoustic signal and the corresponding referencesignal. Eq. (3) establishes a system of linear equations that is solvedusing established iterative methods from the medical imagereconstruction literature. In particular, to mitigate artifacts due tonoise and limited measurements, modern reconstruction methods, inspiredby compressive sampling theory, is utilized for this task.

C. Reconstruction of Ultrasound Reflectivity

Algorithms developed for reconstructing the 3D distribution of acousticreflectivity of the breast from knowledge of reflected, orbackscattered, ultrasound data are provided. These algorithms aredeveloped within the framework of 3D reflectivity tomography. Inprevious theoretical studies (24,25), identified data redundancies wereidentified and it was demonstrated that accurate images could bereconstructed from backscattered acoustic echo data recorded on asampled hemi-spherical measurement aperture. Based on that work, robustiterative reconstruction algorithms that incorporate the effects of thefinite transducer size and finite sampling effects are developed.

IX. Ultrasound-Assisted OAT Image Reconstruction

In previous studies of OAT it was assumed that the object isacoustically homogeneous, which can limit image resolution.Reconstruction approaches for OAT that can compensate for acousticheterogeneities in the determined SOS distribution via inversion of ageneralized Radon transform (GRT) imaging model are developed. We haveextensive experience with this topic (28). Perturbation theory fortravel times is employed to incorporate higher-order diffraction effectsinto the GRT imaging model (28). This is based on a higher ordergeometrical acoustics generalization of the OAT imaging model that takesinto account the first-order effect in the amplitude of the measuredsignal and second-order perturbation to the travel times thatincorporate the effect of ray bending. Data redundancies are exploitedto demonstrate that the GRT model can be inverted uniquely and stably byuse of only half of the acquired measurement data. Iterativereconstruction approaches that permit explicit control of statisticallycomplementary information that can result in the optimal reduction ofimage variances are developed. Methods based on time-reversal principlesalso are investigated. The effects of imperfect knowledge of theacoustic heterogeneity map also be investigated and robust methodsdeveloped to mitigate them. The development of such methods forcompensating for acoustic attenuation is based on previous studies (26).

X. Optimization of Reconstruction Methods Via Computer-SimulationStudies

Computer-simulation studies are conducted to assess quantitatively theperformance of the developed reconstruction algorithms. Realistic 3Dnumerical breast phantoms (16) are constructed that depict theacoustical and optical absorption properties of breast tissue. By use ofthese phantoms, simulation data is computed by solving the acoustic waveequation using the inventors' existing codes. Standard measures ofphysical image quality such as mean squared error is initially used toguide the development and optimization of the algorithms. The impact ofphysical factors such as stochastic data noise, the finite bandwidth ofthe receiving ultrasound transducers, and the effects of finite samplingis investigated and compensated for. The developed algorithms arefurther refined and evaluated in the experimental studies describedbelow that will quantify task-based image quality measures.

Evaluation Studies I. Evaluation of the Imaging System Using PhysicalPhantoms

The imager and algorithm designs is informed and evaluated throughoutthe project by use of experimental studies that utilizewell-characterized multi-modality phantoms made of either gelatin orpoly(vinyl-chloride) plastisol (PVCP) and accurately mimicking opticaland acoustic properties of the object or tissue of interest using TiO2as an optically scattering substance, various dyes for changing opticalabsorption and polystyrene and glass microspheres for changing acousticproperties of the phantoms.

Ultrasonic and optoacoustic phantoms exist or can readily beconstructed. However, a single phantom that is appropriate forvalidating our dual-modality imaging system does not exist. Specializeddual-modality (US+OAT) phantoms that are well characterized can beconstructed. These phantoms incorporate the optical scattering andabsorption properties as well as the acoustic properties of breasttissue and are based on the inventors' hybrid phantoms for use withultrasound tomography and diffuse optical tomography. Concentration ofplastisol in PVCP was varied to achieve appropriate acoustic properties,for example, SOS, density, or attenuation. The use of glass microbeadsto achieve tissue ultrasonic reflectivity were investigated.

For modeling the appropriate optical properties of breast tissue, i.e.,index of refraction, absorption coefficient, scattering coefficient, andscattering anisotropy, dyes, India ink, and titanium oxide powder wereused. PVCP has been shown to possess both optical and acousticproperties similar to tissue, Indian ink is a common optical absorbingmaterial, TiO2 powder is an established choice for modeling opticalscattering, and small glass beads that are optically transparent isexplored as a means of modeling acoustic attenuation of breast tissue.Blood-filled tumor-like inclusions are developed and use colored polymerthreads are used for modeling microvessels. Ultrasonic and optoacousticmeasurements are conducted to validate the phantoms.

II. Phantom Imaging Studies

Phantoms imaging studies are conducted to validate the imager andalgorithms. Experimental parameters that are varied include the numberof tomographic views acquired and the number of optoacoustic signalsacquired at each transducer location that are averaged to improve SNR.The algorithms described herein are designed to reduce both of thesequantities in order to minimize data-acquisition times. By use ofphantoms that have tumors located at different depths and have differentoptical absorption properties, the sensitivity of the OAT system isquantified. Simplified versions of the phantoms are imaged forcharacterizing the spatially variant spatial resolution (42) and noiseproperties (60) of the reconstructed images. Additional image qualitymetrics employed in the imaging method are described below.

In-Vivo Imaging Studies

In Vivo Imaging System in Subjects with Tumors and Lesions Suspected asMalignant

These in-vivo studies fine-tune the imaging system and imagereconstruction algorithms and quantify breast cancer detectionperformance in a clinical setting. Breast cancer imaging represents thefirst in-vivo human application of multimode ultrasound/optoacoustictomography, and yields preliminary data relevant to an evaluation of itsclinical effectiveness. The system is highly effective for therapymonitoring, since laser optoacoustic functional and molecular imagingcan reveal early physiological changes in blood supply, angiogenesisdensity and other molecular biomarkers.

The patient lies in the prone position on the examination table with thebreast suspended through an opening into an imaging tank filled withsterile warm water based optoacoustic coupling medium. The imagersurrounds the breast and collects the multi-wavelength OAT andultrasound tomography measurement data. The multi-wavelength OATmeasurements are acquired using laser wavelengths of 757 and 1064 nm,which permits differentiation of hypoxic and oxygenated blood. Data isacquired at 800 nm, where hypoxic and oxygenated blood absorb equally,i.e., the isosbestic point, which facilitates image normalization. Theappropriate number of tomographic views to acquire to avoid conspicuousartifacts is based on the numerical and physical phantom studies. Fromthese data, tomographic images representing the SOS, attenuation,reflectivity, and absorbed optical energy density are reconstructedonsite by use of the developed algorithms that are most computationallyefficient. The measurement data is saved and is utilized for additionaloff-site processing by use of advanced image reconstruction algorithmsand is utilized to refine the algorithms and systems provided herein.

Patient Population

The clinical study is performed according to an IRB protocol pendingapproval at MD Anderson Cancer Center. Patients with suspicious breasttumors identified by mammography and confirmed by ultrasound as BIRADS 4and 5 and scheduled for biopsy undergo the multimode laser optoacousticprocedure prior to biopsy. As needed, breast MRI is performed onpatients with ambiguous mammography and ultrasound images. Biopsy servesas the gold standard method to determine the tumor pathology. Patientinformation or other data with identifiers linked to the subjects isremoved from any reports that can be taken outside the clinical Center.

Creation of Composite Multi-Parametric Images

The ultrasound tomography images, for example, SOS, attenuation, orreflectivity, may be fused into a single color-coded composite image.Human perception is not well suited to integrating diagnosticinformation presented in a set of related images viewed in parallel(61-63). It is contemplated that image fusion may facilitate thedetection of breast cancer from the multi-parametric ultrasound imagesby a human observer (20). The imaging systems and methods providedherein are useful for forming a single composite image by use of linear(62) and non-linear (61) mappings of the single-parameter image valuesinto red, green, and blue channels. These mappings can encode as muchinformation as possible to help the expert reader. The evaluationmethodology employed, including intra- and inter-observer analysis, isessentially similar to that employed by Alfano, et al. (64) for amulti-spectral MRI application. A similar methodology is utilized tosummarize information regarding the functional OAT images in a singlecomposite image. Thus, a composite OAT image depicting the total bloodconcentration can be color coded with a color of dominating level ofoxygen saturation, so the radiologist can see the brightness based onthe total blood content where the color tells him/her whether the bloodis hypoxic or normally oxygenated.

As described below, the invention provides a number of advantages anduses, however such advantages and uses are not limited by suchdescription. Embodiments of the present invention are better illustratedwith reference to the Figure(s), however, such reference is not meant tolimit the present invention in any fashion. The embodiments andvariations described in detail herein are to be interpreted by theappended claims and equivalents thereof.

FIGS. 1A-1C illustrate the advantages of LOUIS in detection of breastcancer during examination and shows images of a portion of a humansubject body with a cancerous tumor. In FIG. 1A an ultrasonic reflectionimage shows morphology of the subject body with volume of interest(tumor) based on a signal proportional to a product of density and speedof sound. In FIG. 1B an optoacoustic image shows the tumor based onsignals proportional to concentration of the total hemoglobin in thetumor angiogenesis microvasculature. In FIG. 8C an X-ray mammographyimage of the same breast shows radiological density of the subject bodywith no contrast for the volume of interest that includes tumor. TheX-ray image is inconclusive due to high breast density, but the presenceof a tumor is confirmed by the ultrasound showing breast anatomy withenhanced tissue density in the tumor, and by the optoacoustic imageshowing high concentration of hypoxic blood in the tumor angiogenesisproduced by the combined ultrasonic/optoacoustic system in diagnosticimaging of breast cancer.

FIG. 2 is a photograph of the laser optoacoustic ultrasonic system as afully assembled and operating prototype, demonstrating that thisinvention was reduced to practice. This tomography system has thefollowing components and their technical specifications:

A. Pulsed Laser: Nd:YAG pumped Ti:Sapphire laser, Q-switched with pulseduration of 8 ns; wavelength tenability range—532 nm, 730 nm to 850 nm,1064 nm; pulse energy 120 mJ, pulse repetition rate 10 Hz, capability totoggle 2 wavelengths and tune continuously one wavelength.B. Imaging module: Array of 128 ultrawide band ultrasonic transducersmade of piezocomposite materials, 1×1 mm lateral dimensions, 5 MHzcentral frequency. Minimal detectable pressure by the system is about 1Pa, which allows quantitative measurements of the optical absorptioncoefficient in the biomedical objects with accuracy of better thanμa˜0.01/cm. Three bifurcating fiber bundles with circular inputs andarc-shaped linear outputs that can be inserted in any of the 7 slots ofthe imaging module subject body. Plastic polymer caps cover outputs ofthe fiber bundles. The polymer caps are made transparent foroptoacoustic imaging and black for laser ultrasonic imaging. Computercontrolled rotational motor allows precise rotation and positioning ofthe imaging module around the biomedical object of interest. Typically,the module is rotated to 300 positions with 1.2 deg steps to acquirecomplete set of 3D data. This in turn generates 38400 virtual detectorson the spherical surface using 128 piezoelectric transducers, whichresults in accurate 3D images.C. Electronics: The electronics are composed of 4×32 channel analog lownoise high input impedance amplifier boards and 4×32 channel digitaldata acquisition boards with 12 bit ADCs and reconfigurable FPGAmicroprocessors for signal processing and transfer of the information toa computer for image reconstruction using multicore GPU Fermi videocard. The system is computer controlled with dual core CPU.

FIGS. 3A-3B are illustrations of the imaging modules forthree-dimensional laser optoacoustic ultrasonic imaging system, LOUIS.FIG. 3A depicts a design suitable for optoacoustic imaging with combinedlinear-flat plus arc shaped combined array of ultrasonic transducers.FIG. 3B depicts a design suitable for laser ultrasonic plus laseroptoacoustic imaging with arc-shaped array of ultrasonic transducers.

The imaging module 10 has a housing 1 made of hypo-echoic acousticallyabsorbing and scattering material further electrically shielded withexternal metallization. An array of ultrawide-band ultrasonictransducers 2, optimized as detectors in the frequency range from 120kHz to 12 Mhz, is a combined linear plus arc (J-shaped) array of 96ultrawide-band ultrasonic transducers and arc-shaped array of 128ultrawide-band ultrasonic transducers. A translational X-Y-Z stage 3provides flexibility for accurately placing the volume of interest closeto the focal area of the ultrasound transducer array. A computercontrolled rotational motor 6 allows precise rotational positioning ofthe imaging module relative to the volume of interest within a subjectbody.

Fiber bundles 4 a,b,c,d for optoacoustic illumination optimally are madeof 50 micron diameter glass fibers, about 12 mm diameter circular inputand either flat rectangular outputs in 4 a,b or arc-shaped linearoutputs 4 c,d. These 1-into-2 split bundles are designed withcylindrical lens to produce expanding beam of near-infrared laserillumination of tissue for optoacoustic imaging. Two pairs of bundlesare placed in the imaging module. One pair 4 a,c is placed closer to thedetecting array of ultrasonic transducers for optoacoustic imaging ofthe skin outline in backward mode. The second pair 4 b,d is placedfacing each other, orthogonally to the detecting ultrasonic array andalong the diameter of the imaging module for deep tissue optoacousticimaging in orthogonal mode. Fiber bundle 5 for laser ultrasoundgeneration optimally is made of 50 micron diameter glass fibers, about12 mm diameter rectangular input for laser coupling and 33 outputs.I.e., a 1-into-33 split. each having circular output with a diameter ofabout 1 mm. This fiber bundle illuminates laser ultrasonic sources withshort pulses of a laser operating at high pulse repetition rate of about1280 to 2560 Hz.

With continued reference to FIGS. 3A-3B FIGS. 4A-4C are views of a laserultrasonic emitter. The emitter 15 comprises fiberoptic illuminatorholder 7, which is a plate that holds the outputs 5 a of the fiberbundle 5 and is configured to functionally connect with the imagingmodule 10. The fiber bundle 5 comprises multiple sub-bundle outputs 5 a,optimally about 32 to 64 sub-bundles. The sub-bundles are placed on adiagonal 7 b to connect the top and bottom corners of the laserultrasound emission aperture 7 a. The laser ultrasound emission apertureoptimally has a height greater than the height of the volume of interestin the subject body and the width corresponding to angular aperturegreater than the width of the volume of interest. As an example anaperture of about 90 deg is shown. The range of angular apertures mayvary with design from as small as 60 deg to as large as 150 degdepending on the dimensions of the volume of interest within the subjectbody.

A plurality of laser ultrasonic emitters, represented by 8, arehemispherical objects coated with thin layer of highly opticallyabsorbing material for emission of laser ultrasound. Due to a finitediameter of the laser ultrasonic generator, the layer of the coatingmaterial should be spherically shaped to produce closer to ideal virtualspherical source of laser ultrasound. A plate-holder 9 holds theplurality of laser ultrasonic emitters 8, which are optimally separatedat 9 a from the outputs 5 a of the glass optical fiber bundles 5 inorder to provide non-reverberating Delta pulses of ultrasound inwater-like optoacoustic coupling medium.

FIGS. 5A-5C illustrate advances in generation of short (so called,Delta) ultrasound pulses using lasers. FIGS. 5A-5C demonstrate thatinvented designs of laser ultrasound (LU) emitters produce shortnonreverberating pulses of ultrasound with high amplitude (FIG. 5A) andultrawide frequency spectrum (FIG. 5B). FIG. 5C shows wide directivitydiagram of LU generation provided by a design with hemi-spherical tipsof LU sources, which generated close to ideal ultrasonic waves withspherical wavefront. The design with spherical tips is preferred vssmall flat sources due to wider directivity of the emitted LU. Based onthis design other improved designs have been implemented. Efficiency ofthe designed LU source, LUE=5 [kPa]/[mJ/cm²], and for the optimizedspherical source coated with highly thermally expanding materials LUEcan reach over 100 [kPa/[mJ/cm²].

FIG. 6 is a table of Gruneisen parameters which is proportional to theefficiency of laser generation of ultrasound. Gruneisen parameter arepresented for examples of liquid and solid materials with high thermalexpansion and high speed of sound, which enables high laser ultrasoundefficiency. The most important, however, is that the material will havevery strong optical absorption at the laser wavelength employed forgeneration of ultrasound pulses. Such metals as gold and silver whenmade as thin layers possess plasmon resonance absorption which can beused for the benefit of LU generation. Alternatively, polymers such asPDMS or PMMA can be used for LU generation when colored with stronglyabsorbing molecules or particles.

FIG. 7 depicts a design of a hand-held probe for the 2D tomographysystem. Real-time laser ultrasonic imaging can be performed using aspecially designed imaging module miniaturized as a hand-held probe 20.FIG. 7 shows two 4-6 mm ultrasonic transducers 21 a,b as a portion of alinear array of 128 ultrasonic transducers. Fiberoptic bundle 22 isinserted between the two transducers to deliver laser pulses 22 a to anoptically absorbing layer 23, which generates ultrasound pulses 23 a inresponse to the laser pulses. An acoustic lens 24 focuses the lasergenerated ultrasonic pulses into a thin slice of tissue in the volume ofinterest and also helps to collect reflected ultrasonic pulses also onlyfrom a thin layer of tissue being imaged. The acoustic lens can alsoserve as an ultrasound emitter, if made at least partially frommaterials that possess strong optical absorption and significant thermalexpansion.

FIGS. 8A-8C demonstrate advantages of laser ultrasound pulses comparedwith electrically generated ultrasound pulses. Electrically generatedultrasound pulse (FIG. 8A) produced by a standard commercial ultrasonictransducer is strongly reverberating, so that an envelope of this pulsehas to be used in reconstruction of images in ultrasound tomography. Incontrast, laser generated pulse is non-reverberating and possess highamplitude. One skilled in the art can conclude from FIG. 8A and FIG. 8B,that laser ultrasound tomography can achieve spatial resolution about 3times better than that of ultrasound that employs electrically generatedultrasonic pulses. FIG. 8C shows that ultrasonic frequency spectrum oflaser ultrasound pulse is much wider compared with electricallygenerated ultrasound pulse. The ultrawide spectrum of ultrasonicfrequencies available for laser ultrasound not only result in greaterspatial resolution, but also can be used for ultrasonic spectroscopy asa method of tissues characterization with diagnostic purposes usingultrasonic imaging systems.

FIGS. 9A-9B are examples of spatial resolution achieved in LOUIS-3Dsystem. In FIG. 9A three intersecting horse hairs were imaged as asubject body and the optoacoustic image brightness cross-section ispresented for one of the hairs. The horse hairs had diameters about 100to 150 micron. In FIG. 9B the image brightness shows Gaussian shape withFWHM equal to 300 micron. Such spatial resolution is achieved withdetecting ultrasonic transducer array having sensitivity bandwidth from150 kHz to 5 MHz. The image resolution can be further improved withwidening the bandwidth of ultrasonic transducers, reduction of thetransducer lateral dimensions and more accurate system alignment.

FIGS. 10A-10B illustrate the importance of ultrasonic transducerssensitive within ultrawide-band of ultrasonic frequencies. Optoacousticprofiles detected from an absorbing sphere by ultrasonic transducer madeof PZT—standard relatively narrow band ultrasonic transducer (FIG. 10A)and a new ultrawide-band transducer made of single crystal PMN ceramic(FIG. 10B) are shown. A similar profile was observed from MPT singlecrystal ceramics. The profile in FIG. 10A is strongly reverberating,i.e. distorted, while the profile in FIG. 10B shows N-shapednon-reverberating pulse, which can be used for reconstruction ofquantitatively accurate optoacoustic images of a sphere.

FIGS. 11A-11B are 2D projections of three-dimensional optoacousticimages of a skin outline of mouse subject body in vivo obtained withLOUIS-3D using illumination in backward mode. The laser illuminationwavelength of 532 nm and the methods of signal and image processing werechosen to emphasize the skin surface. Knowledge of the skin outlinepermits separation of the volume inside the imaging module into twodomains: the domain of the subject body and the external domain of theoptoacoustic coupling medium. Since all properties of the couplingmedium are well known, separation of the two domains allows much moreaccurate reconstruction of volumetric optoacoustic and ultrasonic imagesof the subject body.

FIGS. 12A-12B illustrates that images of the speed of sound presentsmorphology with valuable diagnostic information. The image in FIG. 12Arepresents distribution of the speed of sound (SoS or SOS) in a phantomsimulating a breast with tumors. Typically breast tumors have an SoShigher than that of normal breast tissues. The image of ultrasonicattenuation (UA) in FIG. 12B represents morphology with valuablediagnostic information, for example, the attenuation of fat andglandular tissues differ in the breast. In addition to diagnosticinformation, SoS and UA images allow correction of optoacoustic andultrasonic images reconstruction algorithms in heterogeneous tissues. Ina human subject body, anatomical ultrasonic imaging can providemorphology of background tissues, SoS and UA information and shape andstructural features of tumors and blood vessels.

FIG. 13 shows a 2D projection of an optoacoustic image of mouse body.The image demonstrates that anatomical images can be produced byoptoacoustic subsystem of LOUIS. Not only soft tissue organs and largervasculature can be visualized, but also microvasculature of the skin,spine, ribs and joints.

FIG. 14 shows a 2D projection of a 3D LOUIS images of an animal subjectbody vasculature, i.e., angiography. Functional optoacoustic imaging canprovide measurements of [Hb] and [HbO] (and total hematocrit) in tissuesand blood vessels, assessment of heart function and blood flow, andassessment of tumor angiogenesis for diagnostic purposes. Microvesselsas small as 50 micron are visible on LOUIS images due to high contrast(resolution 300 micron). Quantitative accuracy of the absorptioncoefficient was found about 0.1/cm in blood vessel phantoms.

FIG. 15 shows an exemplary optoacoustic image of brain vasculature of alive mouse. This type of imaging is important for detection andcharacterization of stroke and traumatic injury of the brain. Thisembodiment demonstrates capability of IOUIS for molecular imaging usingexogenous contrast agents.

FIGS. 16A-16C shows 2D projections of 3D optoacoustic images of breasttumor receptors visualized using targeted contrast agent based onbioconjugated GNRs. Before injection of the contrast agent, mouse tumor(FIG. 16A) was visualized based on its microvasculature (FIG. 16B).After intravenous injection of gold nanorods (GNR) conjugated withPEG-Herceptin (FIG. 16C), distribution of targeted molecular receptorsof HER2/neu in BT474 breast cancer cells became most contrasted feature.Quantitative information is the primary merit of optoacoustic imagingand can provide absolute values of the optical absorption coefficientand concentration of the most physiologically important molecules in thesubject body.

FIGS. 17A-17B are exemplary 3D laser optoacoustic images of the breastacquired and reconstructed with LOUIS-3D. The laser wavelength used forillumination was 760 nm to emphasize veins and tissues with lowblood-oxygen saturation level in the right breast (FIG. 17A) and 1064 nmto emphasize arteries and oxygenated tissue in the left breast (FIG.17B). Optoacoustic orthogonal mode of illumination was used to acquirethese images. Combination of Ultrasound and Optoacoustic Imaging alsocan be produced by LOUIS. One and the same probe and electronicshardware allows coregistration of ultrasonic and optoacoustic images,yielding complementary biomedical information.

FIG. 18 illustrates the method of optoacoustic image reconstruction withhigh accuracy of quantitative information based on data spacerestoration using curvelet transform followed by image reconstructionusing filtered backprojection. This method is real time imaging methodis equal or even more accurate than iterative methods of optoacousticimage reconstruction.

FIGS. 19A-19B show two optoacoustic images of a mouse vasculaturereconstructed using standard filtered backprojection algorithm, whichproduces significant blurring and distortions (FIG. 19A) and filteredbackprojection algorithm using optoacoustic signals processed withcurvelet deconvolution method of data space restoration, which removessignal distortions associated with imperfection of the system hardwareas well as alterations that occur in the course of propagation throughtissues (FIG. 19B).

FIGS. 20A-20B show two images reconstructed using filteredback-projection algorithm taking entire set of measured optoacousticsignal data (FIG. 20A) and iterative algorithm taking only ¼ portion ofthe set of measured optoacoustic signal data (FIG. 20B). This exampleshows that the number of detecting transducers can be optimized bytrading off small reduction in image quality for significant reductionin the data acquisition time and system cost. Based on the presentdesign of LOUIS-3D and our understanding of iterative algorithms of 3Dimage reconstruction using sparse data, we teach here that LOUIS-3D isable to produce real-time volumetric images, i.e. acquire images withvideo rate of multiple frames per second. One possible design of theimaging module is a sphere sparsely but evenly covered with ultrasonictransducers, e.g. 512 detectors, which can acquire 3D images in onestatic position without any rotation around the object. With more andmore powerful computers in future, it is contemplated thatreconstruction of 3D images, i.e., large volumes with very highresolution. also can be accomplished faster than in 1 second. Thefollowing references are cited herein.

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This invention fulfills a longstanding need in the art for a tomographysystem that provides images based on ultrawide-band non-reverberatinglaser-induced ultrasonic pulsed signals. The system providesquantitative functional and molecular plus anatomical imaging throughcoregistered and mutually informed laser ultrasonic and optoacousticimages. The specifications and embodiments described herein serve toprovide for disclosure of the following specific systems, methods andtheir biomedical applications.

What is claimed is:
 1. A dual-modality imaging system, comprising: a)first subsystem comprising a laser ultrasonic imaging system having anultrawide-band ultrasonic transducer array sensitive within a range ofultrasonic frequencies from 50 KHz to 30 MHz positioned for acousticdetection of transient pressure waves resulting from delivery of thelaser generated ultrasound to a subject body and configured to producetomographic images of a volume of interest in the subject body utilizingparameters comprising one or more of the speed of sound, ultrasoundattenuation or ultrasound backscattering; and b) second subsystemcomprising a laser optoacoustic imaging system having one or more arraysof ultrawide-band ultrasonic transducers sensitive within the range ofultrasonic frequencies from 50 KHz to 30 MHz positioned for acousticdetection of transient pressure waves resulting from delivery of shortlaser pulses of nanoseconds duration for generating optoacoustictomographic images of distribution of the optical absorption coefficientin the subject body utilizing parameters of the absorbed optical energydensity or various quantitative parameters that can be derived from theoptical absorption.
 2. The dual-modality imaging system of claim 1,wherein the images generated by the laser-generated ultrasound aretomographic images of tissue anatomy, morphology and structure.
 3. Thedual-modality imaging system of claim 1, wherein the images generated bythe laser-generated optoacoustics are tomographic images of tissuefunctional molecules of hemoglobin, oxyhemoglobin, water, lipids,proteins.
 4. The dual-modality imaging system of claim 1, wherein theimages generated by the laser-generated optoacoustics are tomographicimages of proteins, nucleic acids, and enzymes comprising tissue ofbiomedical interest targeted with exogenous contrast agents or images ofa spatial distribution of the exogenous contrast agents, said contrastagents increasing contrast or characterizing molecules, cells ortissues.
 5. The dual-modality imaging system of claim 4, wherein saidexogenous contrast agents are optical, optoacoustic, acoustic ultrasonicor dual optoacoustic-ultrasonic contrast agents and said contrast agentsare either molecules or nanoparticles.
 6. The system of claim 5, whereinall images are spatially coregistered or temporally coregistered.
 7. Animaging method for increasing contrast, resolution and accuracy ofquantitative information obtained within a subject, comprising the stepsof: a) producing a laser ultrasound of an outline boundary of a volumeof interest within the subject using the dual-modality imaging system ofclaim 1; b) generating a spatially or temporally coregistered image ofat least one of speed of sound and an image of ultrasonic attenuationwithin the outlined volume boundary from information contained in thelaser ultrasound; and c) generating a spatially or temporallycoregistered optoacoustic image based on absorbed optical energy usingat least one of distribution of the speed of sound and ultrasoundattenuation within the outlined volume boundary.
 8. A laser optoacousticultrasound imaging system (LOUIS), comprising: a) a dual laser sourceswitchable between a laser ultrasonic mode and a laser optoacousticmode, said laser source configured to emit either short optical pulseswith high repetition rate for the illumination of the ultrasonicemitters in the ultrasonic mode or short optical pulses with lowerrepetition rate but higher pulse energy for the illumination of thevolume of interest in the optoacoustic mode; b) an imaging modulecomprising one or more ultrawide-band ultrasonic transducers configuredto detect, through a coupling medium, optoacoustic and ultrasonicsignals propagated as transient pressure waves from said volume ofinterest within a subject body; c) means to rotate and/or translate saidimaging module relative to the volume of interest in the subject body tocreate multiple pressure waves, said means computer controllable ormanually controllable; d) means for processing said detected laseroptoacoustic and laser ultrasonic signals and reconstructing processedsignals into one or more of anatomical and functional/molecular imagesof the volume of interest in the subject body; and e) means fordisplaying the one or more images or superimposed coregistered images ofthe subject body or the volume of interest therein.
 9. The laseroptoacoustic ultrasound imaging system of claim 8, wherein laseroptoacoustic illumination is performed in orthogonal mode, backward modeforward mode relative to the subject body or the volume of interesttherein.
 10. The laser optoacoustic ultrasound imaging system of claim8, wherein laser ultrasonication is performed in transmission or forwardmode or in reflection or backward mode relative to the subject body orthe volume of interest therein or in a combination of said modes. 11.The laser optoacoustic ultrasound imaging system of claim 8, wherein thetransducer array is interchangeable for acquisition of various types ofimages in order to achieve greater contrast, resolution, or quantitativeaccuracy of either optoacoustic or ultrasonic images or both.
 12. Thelaser optoacoustic ultrasound imaging system of claim 8, wherein therotating means is configured to rotate said imaging module, wherein thedetecting array of transducers comprises an arc-shaped array or linearflat array or combination of said array shapes comprising ultrawide-bandultrasonic transducers with wide angular directivity.
 13. The laseroptoacoustic ultrasound imaging system of claim 8, wherein thetranslating means is configured to translate said imaging module,wherein the detecting array of transducers comprises an arc-shaped arrayor linear flat array or combination of said array shapes comprisingfinite size ultrasonic transducers with narrow angular directivity. 14.The laser optoacoustic ultrasound imaging system of claim 8, wherein themeans for processing and reconstructing said detected ultrasonic signalscomprises one or more of: electronic amplifiers with time-gain-controlcircuits; multichannel analog-to-digital-converter with a fieldprogrammable gate array; and imaging module design and tomographyalgorithms configured to reconstruct quantitatively accurate volumetricimages.
 15. The laser optoacoustic ultrasound imaging system of claim 8,wherein said imaging module comprises a hand-held probe configured foracquisition, reconstruction and display of real-time two-dimensional orthree-dimensional images.
 16. A method for imaging a subject's body or avolume of interest within, comprising the steps of: a) positioning thesubject body within or proximate to the imaging module of the laseroptoacoustic ultrasound imaging system of claim 8; b) delivering alaser-generated pulses of ultrasonic energy to a volume of interest inthe subject body; c) detecting the transmitted or reflected ultrasonicpressure waves while measuring one or more parameters comprising adifference between the time of emission and a time of arrival, adifference between emitted amplitude and detected amplitude, and adifference between ultrasonic frequency spectrum of emitted and detectedultrasonic pulses; d) delivering a laser-generated pulse of opticalenergy to a volume of interest in the subject body, wherein the pulse ofoptical energy has a duration shorter than the time of pressure wavepropagation through the distance in the subject body or volume thereofequal to a desired spatial resolution; e) detecting the ultrasonicpressure waves generated through optical absorption inside the subjectbody while measuring one or more parameters comprising a time of arrivalrelative to a time of generation, an amplitude of detected optoacousticsignals, and an ultrasonic frequency spectrum of detected optoacousticsignals; f) scanning the subject body or volume of interest therein witha detecting array of ultrawide-band ultrasonic transducers by repeatingsteps b) to e) at multiple positions around the subject body or volumeof interest while simultaneously scanning the sources of optical energyand sources of ultrasonic energy such that relative position of thedetecting array of ultrasonic transducers and the sources of optical orultrasonic energy can change or remain constant during the scans; g)processing the detected ultrasonic signals to remove distortions ofdetected signals; and h) reconstructing one or more volumetric imagesvia mathematical tomography algorithms using data of the processedsignals.
 17. The method of claim 16, wherein the scanning stepcomprises: a) scanning the whole subject subject body with a first arrayof ultrasonic transducers in a rotational configuration to determine atleast one volume-of-interest and its characteristics related to absorbedoptical energy; b) replacing the first array with a second array ofultrasonic transducers in a translational configuration; and c) scanningthrough said at least one volume-of-interest with a high resolutionsufficient to acquire quantitative information related to distributionand concentration of functional molecules therein.
 18. The method ofclaim 16, wherein the one or more volumetric images arethree-dimensional images of the volume of interest or of the subjectbody, or two-dimensional slices through the three-dimensional volume ofinterest or one-dimensional profiles of molecules of interest within thevolume.
 19. The method of claim 16, wherein at least one volume ofinterest is a tumor, a lymph node, a vascular circulation network, or abrain.
 20. The optoacoustic imaging method of claim 16, wherein the stepof delivering pulsed optical energy is performed at multiple wavelengthsof light, whether in sequence or toggling.